Atomic Force Microscopy (AFM) is a powerful measurement technique for nanoscale science and technology. While it is able to provide high-resolution imaging of biological structures below the optical limit and monitoring of biological system dynamics and processes under physiological conditions, certain limitations still exist in the field of bio-applications. In recent times, the development of one class of SPM technique, scanning ion conductance microscopy (SICM), has overcome these limitations and enabled noninvasive, nanoscale investigation of live cells [1]. SICM applications include imaging of cell topography, monitoring of live cell dynamics, mechanical stimulation of live cells, and surface patterning and so forth. Additional findings on the combination of SICM with other SPM techniques as well as patch clamp electrophysiology and optical techniques such as confocal microscopy and fluorescent microscopy can help building integrated knowledge on the structure and function of live cells.Developing AFM for advanced nanoscale metrology, we separated the z-scanner from the x-y scanner [2]. The separating z-scanner also provides an excellent platform for converging other scanning probe microscope techniques such as Near-field Scanning Optical Microscopes (NSOM), Tip-enhanced Raman Spectroscopy, and Scanning Ion Conductance Microscopy (SICM). The glass micropipette which is used in SICM as the sensitive probe, instead of a silicon-based stylus, can glide over live cells while maintaining an absolute non-contact imaging mode, and its electrochemical current feedback system further enhances biological sample imaging. In addition, the glass micropipette enables us to image and conduct nanometer-size electrophysiology patch clamping, subsequently allowing us to identify the exact location of the target molecular species. Combining confocal fluorescence data to the SICM 3D date using image overlay feature provides even more data about structure of cells related to the membrane. We have separately stained nuclei, actin and membrane of human alveolar basal epithelial cells (A549) using DAPI, FITC, and TRITC, respectively and combined with ICM 3D image reconstruction. Image overlay function fully utilizes the convergence capability of Scanning Probe Microscopy (SPM) and provides users with many new possible applications and new challenges in the growing multi-disciplinary environments.[1] F. Anariba, J. H. Ahn, G.-E. Jung, N.J. Cho, and S.-J. Cho, Mod. Phys. Lett. B 26. 1130003 (2012)[2] J. Kwonn, J. Hong, Y.-S. Kim, D.-Y. Lee, K. Lee, S.-M. Lee and S.-I. Park, Rev. Sci. Instrum. 74. 4378 (2003)

SICM convergence microscopy with confocal fluorescence capability and its future perspectives in cell biology

244th National Meeting (2012)

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